Development of a novel approach for the production of dried genomic DNA for use as standards for qualitative PCR testing of food-borne pathogens

As part of a multi-centre European project, FOOD-PCR, the feasibility of a novel approach for production of dried bacterial DNA that could be used as certified reference materials (CRM) was assessed. Selected strains of Salmonella typhimurium, Listeria monocytogenes, Escherichia coli O157, Campylobacter jejuni and Yersinia enterocolitica were used to produce genomic DNA (gDNA). These preparations gave support to method development for qualitative polymerase chain reaction (PCR) detection methods for food-borne pathogens. Purified gDNA was transformed into stable and dry gDNA by using polypropylene vials as carrier and applying a vacuum-drying technique. The gDNA preparations were shown to be sufficiently stable under ambient transport conditions without cooling and proved to have long-term stability at 5°C of at least 22 months. The dried DNA was easily reconstituted by addition of distilled water then gentle shaking. These studies have shown that production of stable and dry bacterial gDNA material is feasible and could help satisfy the increasing need for certified reference DNA positive control samples in the field of PCR testing for detection and verification of food-borne microbial pathogens.

     

Elemental Selenium Enriched Nanofiber Production

This study aimed to produce electrospun nanofibers from a polyvinyl butyral polymer (PVB) solution enriched with red and grey selenium nanoparticles. Scanning electron microscopic analysis was used to observe the samples, evaluate the fiber diameters, and reveal eventual artifacts in the nanofibrous structure. Average fiber diameter is determined by manually measuring the diameters of randomly selected fibers on scanning electron microscopic (SEM) images. The obtained nanofibers are amorphous with a diameter of approximately 500 nm, a specific surface area of approx. 8 m² g⁻¹, and 5093 km cm⁻³ length. If the red and grey selenium nanoparticles were produced in powder form and suspended to the ethanolic solution of PVB then they were located inside and outside the fiber. When selenium nanoparticles were synthesized in the PVB solution, then they were located only inside the fiber. These nanofiber sheets enriched with selenium nanoparticles could be a good candidate for high-efficiency filter materials and medical applications.     

Changes in the heavy metal contents of soil from the Park Grass Experiment at Rothamsted Experimental Station

The classical Rothamsted Experiments allow to contribute to current research. The heavy metal content of soil and hay samples have been measured with ICP-AES from a control and a fertilized plot of the Park Grass Experiment. Today a difference of 1.2 can be observed in the pH levels of the control and fertilized plots; the pH level of the control plot has decreased about 1.0, that of the fertilized plot about 2.2 as a result of 150 years of acid deposition and fertilization. The changes in the pH levels has caused many changes in the heavy metal content of the top layer of the soil. Using different extraction methods (e.g., ammonium acetate, EDTA, and sequential analysis), the heavy metal content of different fractions have been estimated. In the control plot, the Zn, Cd and Pb content increased. In the fertilized plot, the Pb and Cd contents have also increased because of atmospheric deposition and small amounts of lead in the fertilizer. The available form of Cd and Pb increased in both the control and the fertilized plots.     

Estimation of the minimum uncertainty of DNA concentration in a genetically modified maize sample candidate certified reference material

Homogeneity testing and the determination of minimum sample mass are an important part of the certification of reference materials. The smallest theoretically achievable uncertainty of certified concentration values is limited by the concentration distribution of analyte in the different particle size fractions of powdered biological samples. This might be of special importance if the reference material is prepared by dry mixing, a dilution technique which is used for the production of the new and third generation of genetically modified (GMO) plant certified reference materials. For the production of dry mixed PMON 810 maize reference material a computer program was developed to calculate the theoretically smallest uncertainty for a selected sample intake. This model was used to compare three differently milled maize samples, and the effect of dilution on the uncertainty of the DNA content of GMO maize was estimated as well. In the case of a 50-mg sample mass the lowest achievable standard deviation was 2% for the sample containing 0.1% GMO and the minimum deviation was less than 0.5% for the sample containing 5% GMO. Received: 5 December 2000 / Revised: 14 March 2001 / Accepted: 19 March 2001     

Optimization of Plant Sample Preparation and ICP Parameters for Analysis of Phosphorus

Abstract

Parameters of HNO₃-H₂O₂ wet digestion sample preparation method and inductively coupled plasma atomic emission spectrometer     

Development of a novel approach for the production of dried genomic DNA for use as standards for qualitative PCR testing of food-borne pathogens

As part of a multi-centre European project, FOOD-PCR, the feasibility of a novel approach for production of dried bacterial DNA that could be used as certified reference materials (CRM) was assessed. Selected strains of Salmonella typhimurium, Listeria monocytogenes, Escherichia coli O157, Campylobacter jejuni and Yersinia enterocolitica were used to produce genomic DNA (gDNA). These preparations gave support to method development for qualitative polymerase chain reaction (PCR) detection methods for food-borne pathogens. Purified gDNA was transformed into stable and dry gDNA by using polypropylene vials as carrier and applying a vacuum-drying technique. The gDNA preparations were shown to be sufficiently stable under ambient transport conditions without cooling and proved to have long-term stability at 5°C of at least 22 months. The dried DNA was easily reconstituted by addition of distilled water then gentle shaking. These studies have shown that production of stable and dry bacterial gDNA material is feasible and could help satisfy the increasing need for certified reference DNA positive control samples in the field of PCR testing for detection and verification of food-borne microbial pathogens.     

Estimation of the minimum uncertainty of DNA concentration in a genetically modified maize sample candidate certified reference material

Homogeneity testing and the determination of minimum sample mass are an important part of the certification of reference materials. The smallest theoretically achievable uncertainty of certified concentration values is limited by the concentration distribution of analyte in the different particle size fractions of powdered biological samples. This might be of special importance if the reference material is prepared by dry mixing, a dilution technique which is used for the production of the new and third generation of genetically modified (GMO) plant certified reference materials. For the production of dry mixed PMON 810 maize reference material a computer program was developed to calculate the theoretically smallest uncertainty for a selected sample intake. This model was used to compare three differently milled maize samples, and the effect of dilution on the uncertainty of the DNA content of GMO maize was estimated as well. In the case of a 50-mg sample mass the lowest achievable standard deviation was 2% for the sample containing 0.1% GMO and the minimum deviation was less than 0.5% for the sample containing 5% GMO.     

Changes in the heavy metal contents of soil from the Park Grass Experiment at Rothamsted Experimental Station

The classical Rothamsted Experiments allow to contribute to current research. The heavy metal content of soil and hay samples have been measured with ICP-AES from a control and a fertilized plot of the Park Grass Experiment. Today a difference of 1.2 can be observed in the pH levels of the control and fertilized plots; the pH level of the control plot has decreased about 1.0, that of the fertilized plot about 2.2 as a result of 150 years of acid deposition and fertilization. The changes in the pH levels has caused many changes in the heavy metal content of the top layer of the soil. Using different extraction methods (e.g., ammonium acetate, EDTA, and sequential analysis), the heavy metal content of different fractions have been estimated. In the control plot, the Zn, Cd and Pb content increased. In the fertilized plot, the Pb and Cd contents have also increased because of atmospheric deposition and small amounts of lead in the fertilizer. The available form of Cd and Pb increased in both the control and the fertilized plots.     

Effect of Phosphate Fertilization on Micronutrient Content of Maize Plant

Abstract

The effect of fertilization and irrigation on phosphate and metal content in maize was studied in an arable land plot experiment on fertilization effects. A field plot experiment was carried out at the Látókép Farm 15 km from Debrecen. Each treatment consisted of 46 m² plots, arranged in a randomized block design with four replication, where the basic treatment was fertilization; the additional treatments were crop rotation, irrigation and cultivation. The soil is Calcareous Chernozem with 2.8–3.0% humus content. The depth of the humus layer is 70–90 cm. The N-content and original P-content of the soil is average, but it is rich in K. Besides macroelements, there is no shortage of trace elements. The element content of maize was determined with a Labtam 8440M inductively coupled plasma atomic emission spectrometer after digestion. A relationship between phosphate and microelement contents in plant was studied. A linear relationship between phosphate and magnesium, and some microelement contents can be found. Linear correlation in P and Mg content of maize was found strongest (Fig 1.).